肝肺综合征发病机制与治疗方法的基础实验研究

Hepatopulmonary syndrome (HPS) is a serious extrahepatic complication of chronic liver disease with poor prognosis and high case fatality rate.It remains complicated and unknown.Previous studies showed that CX3CL1 / CX3CR1 pathway played an important role in pathological process of inflammation and vasodilation and regenerative diseases by mediating the biological activities.We have found that in lung tissue of HPS models there is a close relationship in both high expression of CX3CL1/CX3CR1 and accumulation of macrophages which lead to angiogenesis and angiectasis bronchiectasis.The early successes will be maken as a breakthrough point for clarifying the molecular mechanism of HPS.Common bile duct ligation(CBDL) models will be established with CX3CR1GFP/GFP-C57BL/6 mice(homozygous CX3CR1-deficient).Through injecting anti-CX3CR1-neutralizing polyclonal antibody and anti-VEGF-monoclonal antibody into the models,We will further clarify the effects and mechanisms of CX3CL1/CX3CR1 and vascular endothelial growth factor in the occurrence and development of HPS,and explain the regulatory relationship between them,to explore the effective treatment methods.Some significantly different cytokines will be screened from bile and plasma between wistar rat and HPS model.Through benefiting from the combination of experiments in vivo and vitro,we will study the relationship between these cytokines and pathogenesis of HPS,to further fully reveal the pathogeneis.The reason of which CBDL is the only way to establish experimental HPS model could be found in this research.These results could provide some new ideas in exploring the reason of which liver transplant is the only method of curing HPS.

肝肺综合征(HPS)是慢性肝病的严重并发症，预后差，病死率高，发病机制复杂且不明确。肝移植是其唯一有效治疗方法。CX3CL1/CX3CR1介导的生物活性参与了多种血管扩张与再生性疾病的病理过程。课题组研究发现HPS模型肺组织CX3CL1/CX3CR1高表达与巨噬细胞聚集而引发的肺微血管扩张与再生有密切关系。本项目为扩展前期研究成果，用CX3CR1基因缺陷小鼠构建肝硬化模型，予CX3CR1多克隆抗体和VEGF单克隆抗体处理HPS模型，旨在阐明CX3CL1/CX3CR1与VEGF途径在HPS发病机制中的作用和机制，探索二者间调控关系及有效治疗HPS的方法。从模型胆汁与血浆中筛选差异明显的细胞因子，经在体与离体实验，探寻在HPS发生发展中其它细胞因子的作用及机制，进一步全面探索HPS的发生机制，有望发现胆总管结扎术是建立HPS模型唯一方法的原因，为探索肝移植是治愈HPS唯一方法的原因提供新思路。

我们的前期研究发现HPS模型肺组织CX3CL1/CX3CR1高表达与巨噬细胞聚集而引发的肺微血管扩张与再生有密切关系。本项目为扩展前期研究成果，拟通过用CX3CR1基因缺陷小鼠构建肝硬化模型，予CX3CR1多克隆抗体和VEGF单克隆抗体处理HPS模型，并结合体外机制研究进一步探讨CX3CL1/CX3CR1 与VEGF途径在HPS发病机制中的作用和机制，探索二者间调控关系及有效治疗HPS的方法。本课题证实了CX3CR1缺陷可减少VEGF和PDGF的产生，抑制p-AKT和p-ERK的活化，下调iNOS、eNOS和HO-1的表达；同时也证实了，与WT小鼠或WT-CBDL小鼠相比，CX3CR1缺陷显著减轻炎症，对模型小鼠治疗性应用CX3CR1多克隆抗体或者应用基因敲除小鼠采取预防性阻断CX3CR1，减缓了肺间质的增厚、肺泡的塌陷融合等。总而言之，本研究实验的证据支持CX3CR1通过抑制AKT/ERK信号通路，调节NO/NOS释放，参与实验性HPS小鼠肺血管生成。本研究的结果有望助于为澄清病理机制和发现未来HPS的治疗策略提供潜在的见解。